Pink water is wastewater generated from the load-assemble-pack (LAP) operations at U. S. Army Ammunition Plants (AAPs). Constituents of pink water include 2,4,6 trinitrotoluene (TNT), trimethylenetrinitramine (RDX) and cyclotetramethylenetetramine (HMX). Carbon sorption is the conventional method for treating this LAP waste. Using granulated activated carbon (GAC) filters, the waste is passed through the GAC removing the explosive constituents by sorption onto the carbon. This method is non-destructive, i.e., the sorbed molecules of contaminant remain intact chemically. Thus, the process generates spent contaminant-laden GAC filters that require further treatment, to include regeneration of the carbon filter for re-use or safe disposal at the end of the filter's useful life. The U. S. military and its contractors generate a substantial amount of spent GAC from pink water treatment and would save considerable resources by replacing the GAC filtration process with a process that actually destroys or neutralizes energetic contaminants. An alternative to GAC is microbial treatment.
Because TNT has a high electron deficiency, any initial microbial transformation is reductive. Vorbeck, C., et al., Initial Reductive Reactions in Aerobic Microbial Metabolism of 2,4,6-Trinitrotoluene, Appl. Environ. Microbiol. 64, 246-252, 1998. Treating TNT waste streams under aerobic conditions produces a variety of partially reduced TNT daughter products. Vorbeck et al. 1998; Pasti-Grigsby, et al., Transformation of 2,4,6-trinitrotoluene (TNT) by Actinomycetes Isolated from TNT- Contaminated and Uncontaminated Environments, Appl. Environ. Microbiol. 62, 1140-1123, 1996; Fiorella, P. D. and Spain, J. C., Transformation of 2,4,6-Trinitrotoluene by Pseudomonas pseudoalcaligenes JS52, Appl. Environ. Microbiol. 63, 2007-2015, 1997. These daughter products converge TNT to aminodinitrotoluenes, diaminonitrotoluenes, tetranitroazoxytoluenes and combinations thereof. Roberts, D. J., et al., Optimization of an Aerobic Polishing Stage to Complete the Anaerobic Treatment of Munitions-Contaminated Soils, Environ. Sci. Technol. 30, 2021-2026, 1996. Under aerobic conditions, these metabolites are resistant to further degradation and are therefore considered dead-end products. Vorbeck et al., 1998. Anaerobic conditions are required for the complete reduction of TNT to triaminotoluene (TAT). Funk, S. B., et al, Initial-phase Optimization for Bioremediation of Munition Compound-Contaminated Soils, Appl. Environ. Microbiol. 59, 2171-2177, 1993; Boopathy, R. and Kulpa, C. F., Trinitrotoluene (TNT) as a Sole Nitrogen Source for a Sulfate Reducing Bacterium Desulfovibrio sp. (B Strain) Isolated from an Anaerobic Digester, Current Microbiol., 25:235-241, 1992; McCormick, N. G., et al., Microbial Transformation of 2,4,6-Trinitrotoluene and Other Nitroaromatic Compounds, Appl. Environ. Microbiol. 31, 949-958, 1976; Preuss, A., et al., Anaerobic Transformation of 2,4,6-trinitrotoluene (TNT), Arch. Microbiol. 159, 345-353, 1993; Hwang, P., et al., Transformation of TNT to Triaminotoluene by Mixed Cultures Incubated under Methanogenic Conditions, Environ. Toxic. Chem. 19, 836-841, 2000. The fate of TAT is largely unknown, although phenolic products of TAT hydrolysis and an adduct of TAT have been identified, apparently formed by the condensation of TAT and pyruvic aldehyde. Lewis et al., 1996; Lewis, T. A., et al., Products of Anaerobic 2,4,6-Trinitrotoluene (TNT) Transformation by Clostridium bifermentans, Appl. Environ. Microbiol. 62, 4669-4674, 1996.
RDX is reported to be more easily biodegraded under anaerobic, rather than aerobic conditions. Funk et al., 1993; Kitts, C. L. et al., Isolation of Three Hexahydro-1,3,5-Trinitro-1,3,5-Triazine-Degrading Species from the Family Entrobacteriaceae from Nitramine Explosive-Contaminated Soil, Appl. Environ. Microbiol., 60:4608-4711, 1994; McCormick, N. G., et al., Biodegradation of hexahydro-1,3,5-trinitro-1,3,5-triazine, Appl. Environ. Microbiol. 42, 817-823, 1981; Roberts et al., 1996. One of the few exceptions includes RDX biodegradation by a white rot fungus. Fernando and Aust, Biodegradation of Munition Waste, TNT (2,4,6-trinitrotoluene), and RDX (hexahydro-1,3,5-trinitro-1,3,5-Triazine) by Phanerochaete chrysosporium, Emerging Technologies in Hazardous Waste Management II, ACS Symposium Series, 468, 214-232, 1991. Another exception is degradation by the bacterium Stenotrophomonas maltophilia PB1 when using RDX as the sole source of nitrogen. Binks, P. R., et al., Degradation of hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX) by Stenotrophomonas maltophilia PB1, Appl. Environ. Microbiol., 61, pp 1318-1322, 1995. A third exception occurs during composting of explosives-contaminated soil. Williams, R. T., et al., Composting of Explosives and Propellant Contaminated Soils Under Thermophilic and Mesophilic Conditions, J. Indus. Microbiol., 9, 137-144, 1992.
Most of the studies demonstrating RDX biodegradation under anaerobic conditions were conducted under poorly defined conditions where the electron donor and acceptors were not firmly established. For example, the bacterial cultures were grown in nutrient broth. McCormick et al., 1981. They were also grown in yeast extract. Kitts et al., 1994. A third example involves growth in Brain Hear Infusion media. Regan, K. M., and R. L. Crawford, Characterization of Clostridium bifermentans and its Biotransformation of 2,4,6-trinitrotoluene (TNT) and 1,3,5-triaza-1,3,5-trinitrocyclohexane (RDX), Biotechnology Letters, 16, 1081-1086, 1994. In the latter two cases, RDX biodegradation was carried out by pure cultures of bacteria isolated from explosives-contaminated soil. In the former case, the nutrient broth was inoculated with organisms from activated sludge.
Anaerobic conditions are required for complete explosives degradation, but little is known about the bacteria carrying out the transformation or the environmental conditions that would further enhance the degradation activity. A preferred embodiment of this invention was motivated by the explosives biodegradation research results at the U.S. Army Engineer Research & Development Center (ERDC), Construction Engineering Research Laboratory (CERL). Adrian, N. R., and Lowder, A., Biodegradation of RDX and HMX by a Methanogenic Enrichment Culture, P 1-6, in B. C. Alleman (ed.) Bioremediation of Nitroaromatic and Haloaromatic Compounds, Battelle Press, Ohio, 1999; Adrian, N. R. and Sutherland, K., RDX Biodegradation by a Methanogenic Enrichment Culture Obtained from an Explosives Manufacturing Wastewater Treatment Plant, US Army CERL Technical Report 99/15, ADA 360452, 1998; Adrian, N. R. and Sutherland, K., Biodegradation of RDX by a Methanogenic Enrichment Culture Obtained from an Explosive Manufacturing Wastewater Treatment Plant, Abst. Ann. Meet. Am. Soc. Microbiology, Q-359, Miami, Fla., 1997. The authors investigated biodegradation of energetic compounds under anaerobic conditions using ethanol as the sole electron donor. They found that the inhibition of the methanogens did not affect the degradation of RDX and did not even appear to be involved in the degradation of RDX. Adrian and Sutherland, 1998. Adding RDX to the mixed culture did, however, inhibit methane generation. Adrian and Lowder, 1999. That same year experiments demonstrated the use of H2 and the involvement of acetogenic bacteria in carrying out explosives degradation. Further evidence indicates that the acetogens are the major metabolic group in the mixed culture that degrades TNT, RDX, and HMX. Arnett, C. and Adrian, N. R., Isolation of a RDX-Degrading Acetogenic Bacterium from a Methanogenic Culture that Degrades RDX, HMX, and TNT, Second Int. Sym. Biodegradation of Nitroaromatic Compounds and Explosives, Leesburg, Va., 1999.
In a parallel effort to the above microbiological research, the ERDC/CERL conducted engineering evaluation of treatment of energetic compounds using biotechnology. A granular activated carbon fluidized bed reactor (GAC-FBR) was demonstrated to biologically degrade energetic compounds under anaerobic conditions at bench and pilot scales. Berchtold, S. R., et al., Treatment of 2,4-Dinitrotoluene Using a Two Stage System: Fluidized-Bed Anaerobic GAC Reactors and Aerobic Activated Sludge Reactors, Water Env. Res., 67, 1081-1091, 1995; Maloney, S. W., et al., Anaerobic Fluidized Bed Treatment of Propellant Wastewater, Water Env. Res., 70, n1, p.52, 1998. These investigations confirmed other work that found that, based on mass balance, end products from TNT degradation included nitrate and biomass in the effluent. VanderLoop, S. L., et al., Biotransformation Of 2,4-Dinitrotoluene under Different Electron Acceptor Conditions, Water Research, 33, n5, p. 1287, 1999; VanderLoop, S. L., et al., Two-Stage Biotransformation of 2,4,6-trinitrotoluene under Nitrogen-Rich and Nitrogen-Limiting Conditions, Water Env. Res., 70, n2, p. 189, 1998. Successful GAC-FBR treatment of TNT was demonstrated at McAlester AAP, Oklahoma. Maloney, S. W., et al., Anaerobic Treatment of Pink water in a Fluidized Bed Reactor Containing GAC, Journal of the American Institute of Chemical Engineers, accepted for publication, 2001.
Certain U.S. patents relate to bioremediation of explosive contaminated soils and groundwater. U.S. Pat. No. 6,066,772, Treatment of TNT-contaminated soil to Hater et al., May 23, 2000 and U.S. Pat. No. 5,998,199, Compost Decontamination of Soil Contaminated with TNT, HMX and RDX with Aerobic and Anaerobic Microorgdnisms, to Moser and Gray, Dec. 7, 1999, use natural organisms to degrade the explosives contaminated soil in anaerobic compost following aerobic compost processes. To achieve an anaerobic condition, at least one oxidizable carbon source is added.
U.S. Pat. No. 5,387,271, Feb. 7, 1995; U.S. Pat. No. 5,616,162, Apr. 1, 1997; and U.S. Pat. No. 6,084,150, Jul. 4, 2000, each issued to Crawford et al., and each entitled Biological System for Degrading Nitroaromatics in Water and Soils, use two-step anaerobic treatment to decontaminate soil and water containing nitroaromatics. The first step is fermentation of a carbohydrate such as starch by facultative microorganisms, thus exhausting oxygen and insuring anaerobic conditions. The second step uses anaerobic microorganisms to destroy energetic compounds, while using a carbohydrate for the carbon and energy source.
U.S. Pat. No. 6,051,420, Method for the Decontamination of Soil Organic Explosives, to Radtke and Roberto, May 20, 1998, describes a method for decontaminating soil in situ by first providing an organic solvent to dissolve chunks of explosives, thus facilitating bioremediation of the smaller particles by bacteria while further being aided by the addition of an organic nutrient to the treated soil.
Each of these methods involves adding a carbon source and multiple stages that may require extensive time to complete. A need for a treatment plan that does not require the addition of an expensive carbon source nor require an extended treatment period is met by a preferred embodiment of this invention.